Comparison of Effects Upon Intraocular Inflammation and Murine Aqueous Humor Dynamics Parameters of Intravitreal Injection of Ad5.CMV.hTGFβ₂^C226/228S and LV.hTGFβ₂^C226/228S

Abstract

Purpose : Intravitreal (IVT) injection in mice of adenoviral 5 (Ad5) vector carrying the hTGFβ₂^C226/228S gene leads to elevation of intraocular pressure (IOP) by recapitulating changes to the trabecular meshwork seen in primary open-angle glaucoma (POAG). But mild to moderate inflammation often results. Lentiviral (LV) vectors can achieve similar gene transduction but without causing inflammation. Here we compared, in mice, the effects of IVT injection of Ad5.CMV.hTGFβ₂^C226/228S with LV.CMV.hTGFβ₂^C226/228S on IOP elevation and other parameters of aqueous humor dynamics (AHD), specifically: aqueous humor flow rate (Fin), total aqueous outflow facility (Ctot), uveoscleral outflow rate (Fu), and episcleral venous pressure (EVP).

Methods : C57BL/6J mice (♀, 3-6 months, 20-30g) were utilized. Conscious baseline (BL) IOP measurements (TonoLab rebound tonometer) and ophthalmoscopic examinations were made in restrained animals. Animals were then divided into 2 cohorts (N=5/cohort), anesthetized with inhalation of isoflurane (2.5%) and O_2(g) (0.8L/min), and given a single IVT injection (2µL) in one eye of a suspension of either: Ad5.CMV.hTGFβ₂^C226/228S (2×10⁷pfu) or LV.CMV.hTGFβ₂^C226/228S (2×10⁶pfu). Contralateral eyes were injected (2μL) respectively with Ad5.CMV.Null or LV.CMV.Null. IOP measurements and ophthalmoscopic observations were made OU once per week commencing 1 week following injection. When IOP was elevated, animals were anesthetized (ketamine (100mg/kg)/xylazine (10mg/kg)) and AHD parameters were determined (constant flow infusion).

Results : Ad5.CMV.hTGFβ₂^C226/228S injected eyes exhibited a mild to moderate tendency towards iridial hyperemia and lenticular cataract. A similar but less severe trend was seen in Ad5.Null-injected eyes. No such trend was seen in any LV-injected eyes. Ad5.Null or LV.Null-injected eyes showed no significant change in IOP from BL. IOP and AHD parameters are shown in the Table (below).

Conclusions : Both Ad5 and LV vectors are capable of generating an elevated IOP model in mice. However, while the Ad5 vector promotes intraocular inflammation, the LV vector does not. The LV vector takes a significantly longer time to generate its effects, and the IOP elevation is greater (73% over BL for LV @7 weeks vs. 48% over BL for Ad5 @2 weeks). Both vectors promote an increase in EVP.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.